Bacteriophage and host mutants causing the rolling‐circle λ DNA replication early after infection

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Abstract
There are two modes of bacteriophage λ DNA replication during its lytic development in Escherichia coli cells. The circle-to-circle (θ) replication predominates at early stages of the phage growth, whereas rolling-circle (σ) replication occurs late after infection to produce long concatemers that serve as substrates for packaging of λ DNA into phage proheads. The mechanism regulating the switch from θ to σ replication remains unknown. Our previous genetic studies indicated that the bacteriophage λ Pts1πA66 mutant cannot replicate at 43°C in the wild-type E. coli host, but it can replicate in the dnaA46(ts) mutant. Density shift experiments suggested that the parental DNA molecules of the infecting phage enter σ replication. Here, using electron microscopy, we demonstrate that as soon as 5 min after infection of the dnaA46(ts) mutant by the λPts1πA66 phage at 43°C, the σ replication intermediates are highly predominant over θ replication intermediates, contrary to the wild-type conditions (wild-type bacteria infected with the λP+ phage). The initiation of replication of the λPts1πA66 mutant at 43°C was strongly inhibited in the dnaA+ host, as demonstrated by electron microscopy and by pulse-labeling of the phage-derived plasmid replicon. Implications for the mechanism of the regulation of the switch from θ to σ replication mode are discussed